Monoclonal antibodies (mAbs) have been employed as targeting biomolecules for the delivery of radionuclides into tumor cells in radioimmunotherapy (RIT). Numerous clinical trials have been performed to validate this modality of cancer therapy (see, for example, Parker et al., Pure Appl. Chem., 63, 427-463 (1991); Chakrabarti et al., J. Nuc. Med., 3.7, 1384-1388 (1996); Sharkey et al., Cancer Res., 48, 3270-3275 (1988); Sharkey et al., Cancer Res., 48, 3270-3275 (1988); and Lee et al., Cancer Res., 50, 4546-4551 (1990)). Several useful β−-emitting radionuclides, including 131I, 90Y, 177Lu, 153Sm, 213Bi, 212Bi, 212Pb and 225Ac, have been employed for labeling mAbs for RIT applications (Denardo et al., Cancer, 73, 1012-1022 (1994); Scott et al., Cancer, 73, 993-998 (1994); Schlom et al., Cancer Res., 51, 2889-96 (1991)).
While one critical variable that influences the effectiveness of RIT is the choice of the radionuclide and its associated emission characteristics, an equally important aspect is the choice of the chemical means by which the radionuclide is bound to the protein. For RIT applications, 90Y or 177Lu must be linked as the metal complex to a monoclonal antibody (mAb) or immunoprotein via a suitable bifunctional chelating agent, wherein that complex must be adequately thermodynamically and kinetically stable to minimize release of the isotope in order to minimize toxicity in vivo (Gansow et al., Nucl. Med Biol., 18, 369-381 (1991)).
The pure β−-emitting radionuclide 90Y (Emax=2.28 MeV; t1/2=64.1 h) has been extensively studied in RIT due to its physical properties (see, for example, Martell et al., Critical Stability Constants, Vol. 1: Amino Acids. Plenum Press: New York, 1974; pp. 281-284; Wessels et al., Med Phys., 11, 638-645 (1984); Chinol et al., J. Nucl. Med., 28, 1465-1470 (1987); and Mausner et al., Med Phys., 20, 503-509 (1993)). The macrocyclic chelating agent 1,4,7,10-tetraazacyclododecane-N,N′,N″,N′″-tetraacetic acid (“DOTA”)
is well-known to be an effective chelator of Y(III) and lanthanides. Numerous bifunctional analogs of DOTA suitable for protein conjugation have been reported in the literature (Szilágyi et al., Inorg. Chim. Acta., 298, 226-234 (2000); Kodama et al., Inorg. Chem., 30, 1270-1273 (1991); Kasprzyk et al., Inorg Chem., 21, 3349-3352 (1982); Cox et al., J. Chem. Soc. Perkin Trans. 1, 2567-2576 (1990); Kline et al., Bioconjugate Chem., 2, 26-31 (1991); and McCall et al., Bioconjugate Chem., 1, 222-226 (1991)).
However, the formation kinetics associated with the DOTA chelating agent also has been found to be less than optimal, requiring either lengthy radiolabeling protocols and/or the use of elevated temperatures to achieve acceptable yields and specific activities. (Ruegg et al., Cancer Res., 50, 4221-4226 (1990); Lewis et al., Bioconjugate Chem. 5, 565-576 (1994)) Alternate approaches for using DOTA have resulted in the development of numerous derivatives wherein modifications of the DOTA framework have been explored to address this deficiency. This has been pursued by either the addition of external chelating moieties (Takenouchi et al., J. Org. Chem., 58, 6895-6899 (1993)), conversion of one of the carboxylates to an amide for conjugation purposes (Lewis et al., Bioconjugate Chem., 5, 565-576 (1994); Lewis et al., Bioconjugate Chem., 12, 320-324 (2001); and Peterson et al., Bioconjugate Chem., 10, 316-320 (1999)), or altering the carbon chain length of the carboxylate (Keire et al., Inorg Chem., 40, 4310-4318 (2001)). While all of these investigations have met with varying levels of success with actual use, resolution of inherently slow formation kinetics and radiolabeling inefficiency remains.
For example, several bifunctional derivatives of DOTA have been synthesized for radiolabeling proteins, including 2-(p-isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (C-DOTA), and 1,4,7,10-tetraaza-N-(1-carboxy-3-(4-nitrophenyl)propyl)-N′,N″,N′″-tris(acetic acid) cyclododecane (PA-DOTA)

One aspect that all of these previously reported derivatives have in common is that the tetraaza ring was retained without any modification. There have been numerous detailed studies of the mechanism of metal ion complex formation with DOTA, as well as with amido and phosphorus analogues (Forsberg et al., Inorg. Chem., 34, 3705-3715 (1995); and Howard et al., Chem. Commun., 1381-1382 (1998)). These studies generally propose a two-step mechanism of electrostatic capture of the metal, followed by encapsulation, during which there is deprotonation of the amines and an associated energy cost due to arrangement of the carboxylates in the proper geometries for metal binding (Howard et al. (1998), supra). One aspect of this process also includes arranging the 12-membered ring into the proper spatial geometry, a process that also has an associated energy cost. The final geometry of the lanthanide DOTA complexes has been well reported (Forsberg et al., (1995), supra; and Howard et al. (1998), supra). Forsberg and co-workers have reported on the preferential ring geometry of these complexes via modeling the tetra-amido DOTA complexes (Forsberg et al., (1995), supra).
The value of having a ligand conjugate to chelate metal ions for therapeutic, diagnostic, or other uses is of commercial importance. This commercial importance is due to the fact that many metal ions have desirable characteristics for these various uses, but the delivery systems for the metal ions lack specificity to target cells or do not adequately bind the metal ions.
Therefore, there is still a need for compounds that possess complex stability comparable to that of DOTA and increased stability in vitro and in vivo. The invention provides such compounds, complexes and compositions thereof and methods related thereto. These and other objects and advantages, as well as additional inventive features, will be apparent from the description of the invention provided herein.